The long-term objectives of the proposed studies are to define and characterize developmental changes (or stage-specific markers) in the mouse mammary epithelium associated with differentiation from the virgin to the lactating state. The developing mammary ductal epithelium is structurally and functionally heterogeneous: there are three physiologically distinct subpopulations of epithelial cells, ductal epithelial, alveolar (secretory) epithelial and myoepithelial. The specific aims will be to: 1) purify and characterize mammary epithelial membrane glycoproteins or proteins that distinguish the three differentiation states of the mouse mammary epithelium (virgin, pregnant and lactating); 2) produce monoclonal and/or monspecific antibodies to purified mammary epithelin-specific antigens; 3) determine the in vivo cellular distribution of the antigens in mammary tissue sections (virgin, pregnant and lactating) by immunocytochemistry; 4) determine differences in antigen marker expression with different substratum, the cellular distribution (membrane versus cytoplasmic) and the role of lactogenic hormones; 5) correlation of cellular morphogenesis and immunocytochemistry for new markers with already established antigen markers of mammary epithelial cells (i.e., cytokeratins, casein, theioesterase II and milk fat globule antigens). Mammary tissues will be obtained from all developmental stages of mouse mammary gland (i.e., virgin leads to pregnant leads to lactating leads to involuting). Dissociated cells released by enzyme treatment will be radiolabeled (125I surface labeling, 3H- or 14C-fucose metabolic labeling) and standard biochemical techniques of detergent cell lysis and limited surface proteolysis will be used to obtain cellular extracts. Mammary cell extracts will be analyzed and purified using SDS-PAGE, immunoprecipitation, immunoblotting, lectin affinity chromatography, and immunoadsorbent chromatography. Monoclonal and/or monospecific antibodies will be produced to purified murine mammary epithelial markers and used for immunocytochemistry with undissociated mammary tissue sections and cell cultures to characterize mammary epithelial subtypes (i.e., ductal, alveolar (secretory) and myoepithelial). The formation of cancers in the mammary gland of animals and humans appears to be influenced by developmental changes occurring in the gland after birth and thus greater knowledge of mammary epithelial cell morphogenesis will be helpful for understanding carcinogenesis.